The microbial etiology of dental caries continues to be researched and debated. While S. mutans is recognized as a prominent cariogenic species, caries - even severe caries - have been observed in the absence of detectable levels of S. mutans. It has been postulated that certain strains among primary plaque- colonizing species of streptococci may represent an alternative etiology of dental decay. These strains have been designated low-pH, non-mutans-streptococci (non-MS) variants. They are significantly more acidogenic and aciduric than is typical for their species and rival the acid-related properties of . mutans. Data from a handful of cross-sectional studies supports the hypothesized role for the low-pH, non-MS in caries etiology. In this application we propose to further test the hypothesis that low-pH, non-MS strains have a potential role in caries etiology by taking advantage of an ongoing study, the Iowa Fluoride Study, that has collected and banked site-specific samples from the occlusal surfaces of second molars from subjects enrolled long-term within the study. In particular, we have samples from identical sites within subjects at ages 13 and 17. Some of these sites show incipient decay (white-spot lesions) at age 17 after being clinically sound at age 13. We propose to: 1) Isolate non-MS strains of primary colonizing streptococci from second molar occulsal samples, determine the diversity of genotypes present, and the diversity of acid-related phenotypes for those strains; and 2) Correlate the acid-related phenotypes with the site-specific caries experience of the subjects from whom they were isolated. The study design will allow longitudinal and cross-sectional data analyses that represent a unique approach to testing this important hypothesis.